Cryopreservation has been an effective method for long-term storage of biological material such as islets. Long-term storage of cells and tissue for use in clinical transplantation is based on the inherent need to collect adequate donor cells or tissue and to have this available at times that are suitable for transplantation into a patient. For example, since current methods to isolate and purify sufficient numbers of islets of Langerhans from a human pancreas are limited and multiple donors are required for successful reversal of insulin-dependent diabetes mellitus, long-term storage allows the collection of adequate quantities of islets for subsequent transplantation. Also, with long-term storage, the recipient pool can be extended to include patients in other medical centers.
Current islet cryopreservation protocols generally are based on protocols originally developed for the cryopreservation of rodent islets. See Rajotte et al., "Viability Studies on Frozen-Thawed Rat Islets of Langerhans," Cryobiology, 14:116-120 (1977) (which is hereby incorporated herein by reference). These protocols use multiple glass freezing tubes and aliquot small quantities of islets per individual glass freezing tube. When these freeze-thaw protocols are expanded to large animals and humans, greater numbers of freezing tubes are needed to accommodate the increased number of islets being frozen. This may amount to 30-40 tubes for a typical human islet isolation. Also, the cryopreservation of large numbers of freezing tubes is labor-intensive and has an increased potential for microbial contamination. In addition, seepage of liquid nitrogen into the glass tubes during low-temperature storage is a potential hazard since the tubes most likely will explode if thawed without expelling the liquid nitrogen.
Thus, there is a need to provide a more effective method for cryopreservation of biological material in bulk (e.g., entire preparations of isolated islets) efficiently and with minimal risk of contamination or loss.
There is also a need to provide a more effective method for stabilizing, storing, and retaining the viability of biological material for transplantation, since multiple donors are usually required for the collection of materials, such as islets in adequate quantities. Collecting these materials over time requires proper storage and then culturing; therefore, improvements in the processes are needed whereby the viability of the materials is retained.